| Autor: |
Tsyrulnyk, A. O., Fedorovych, D. V., Sobchuk, S. M., Dmytruk, K. V., Sibirny, A. A. |
| Předmět: |
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| Zdroj: |
Microbiological Journal / Mikrobiolohichnyi Zhurnal; Sep/Oct2021, Vol. 83 Issue 5, p3-10, 8p |
| Abstrakt: |
Riboflavin (vitamin B2) is required for synthesis of the flavin coenzymes: riboflavin-5'-phosphate (flavinmononucleotide) and flavin adenine dinucleotide. Riboflavin is important biotechnological commodity withannual market around 250 million US dollars. It is mostly used as component of feed premixes for animals(80%), in food industry as food colorant, in medicine and component of multivitamin mixtures and asdrug for treatment of some diseases. Over the past two decades, the microbial production of riboflavin byfermentation completely replaces the chemical synthetic route. The main producers of riboflavin in industryare engineered strains of the bacterium Bacillus subtilis and of the mycelial fungus Ashbya gossypii. Flavinogenic yeast Candida famata has great biosynthetic potential. Using combination of classicalselection and metabolic engineering (overexpression of SEF1, RIB1 and RIB7 genes coding the positiveregulator, the first and the last structural enzymes of riboflavin synthesis) resulted in the construction of genetically stable strain of C. famata that produces 16 gram of riboflavin per liter in bioreactor. However, the productivity of riboflavin biosynthesis remains still insufficient for industrial production of thisvitamin. Studies of transcriptional regulation of genes involved in riboflavin synthesis and using of strongpromoters of C. famata for construction of efficient producers of vitamin B2 are areas of both scientificand industrial interest. Aim. The aim of the current work was to improve riboflavin oversynthesis by theavailable C. famata strains in synthetic and natural lactose-containing media. Methods. The plasmidDNA isolation, restriction, ligation, electrophoresis in agarose gel, electrotransformation, and PCR werecarried out by the standard methods. Riboflavin was assayed fluorometrically using solution of syntheticriboflavin as a standard. The cultivation of yeasts was carried out in YNB or YPD media containingdifferent source of carbon and on whey. Results. The strains of C. famata expressed additional copy of central regulatory gene SEF1 under control of the promoter of LAC4 gene (coding for β-galactosidase)C. famata were constructed. The influence of SEF1 gene expression under control of lactose induciblepromoter of CfLAC4 gene on riboflavin production was studied. It was shown that the C. famata strainscontaining "pLAC4_cf-SEF1_cf" expression cassette revealed 1.6-2.1-fold increase in riboflavin yieldon lactose when compared to the parental strain. The riboflavin production constructed strains on wheyreached 1.69 gram per liter in flask batch culture. Conclusions. The constructed strains containingadditional copy of SEF1 gene under the control of LAC4 promoter is a perfect platform for developmentof industrial riboflavin production on by-product of dairy industry, whey. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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